Thiorphan, or (2-mercaptomethyl-3-phenyl-propionylamino)-acetic acid, is a thiol-containing drug that has been clinically administered as its S-acetyl O-benzyl prodrug racecadotril (also known as acetorphan) for the treatment of diarrhea (Matheson A J, Noble S. Drugs 2000; 59: 829-835; Huijghebaert et al., Dig Dis Sci 2003; 48: 239-245). Thiorphan shows low-nanomolar inhibitory activity against neutral endopeptidase (NEP or enkephalinase), a zinc-metallopeptidase widely distributed in peripheral tissues and in the brain, whose biological functions include catabolism of the opioid peptides (Rogues et al., Nature 1980; 288: 286-288; Lecomte et al., J Pharmacol Exp Ther 1986; 237: 937-944).
Enkephalinase inhibitors have received comprehensive studies for their potential therapeutic applications, namely in CNS and digestive tract diseases (Beamont A, Fournie-Zalusk M-C, Rogues B P. In Zinc Metalloproteases in Health and Disease, Hooper N M (ed.) Talor & Francis: London, UK, 1996; 105-129.). Tritium labelled thiorphan (Pollard et al., Eur J Pharmacol 1987; 133: 155-164, De la Baume et al., Eur J Pharmacol 1988; 149: 121-129) and acetorphan (De la Baume et al., J Pharmacol Exp Ther 1988; 247: 653-660) have been used as radioactive probes for characterization of enkephalinase.
Tritium and deuterium labelling of thiorphan is difficult due to the presence of a mercapto functional group, which is troublesome for reactions often used in aromatic tritiation, such as halogenation and catalytic halogen-tritium displacement.
There are two types of tritium labelled thiorphan known in the literature: one with the tritium labelling on the glycine methylene portion (formula 1)
which is prepared using commercially available [3H]-glycine as tritium source (Pollard et al., Eur J Pharmacol 1987; 133: 155-164, De la Baume et al., Eur J Pharmacol 1988; 149: 121-129), and the other with tritium labeling on the phenyl ring portion (formula 2)
which is prepared through tritium-halogen exchange reaction via preparation of the corresponding halo-substituted precursor (Fournie-Zaluski et al., Pept Synth Struct Funct: Proc 7th Am Pept Symposium, Rich D H, Gross E (ed). Pierce Chem Co: Rockford, Ill., 1981; 425-428). The synthetic yields for both types of tritium-labeled thiorphan are very low, due to either catalyst poisoning by the divalent sulfur group or to lengthy multi-step synthetic procedures. Direct halogenation of thiorphan or acetorphan leads to sulfur oxidation, forming a disulfide during aromatic iodination with bis(pyridine)iodonium tetrafluoroborate, or forming a sulfonic acid upon bromination. These problems have limited the availability and use of [3H]- and [2H]-thiorphan. Therefore, a new and robust synthesis of [3H]- and [2H]-thiorphan is needed.